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Zhao Y.,Biogas Institute of Ministry of Agriculture
Wei sheng wu xue bao = Acta microbiologica Sinica | Year: 2012

To identify a thermophilic bacterium from horse manure to degrade cellulose efficiently, and to enrich microbial resources producing cellulolytic ethanol by co-culturing with thermophilic ethanol producing bacterium. We used Hungate anaerobic technique to isolate a strain named as HCp from horse manure mixed culture; its phylogeny was identified through 16S rDNA sequencing. Enzymatic assays were determined using DNS method. The isolated HCp cells were straight with rods size of(0.35-0.50) microm x (2.42-6.40) microm, in the form of single or paring. This strain belongs to a strictly anaerobic Gram-negative bacterium, it is able to form spores, shows motile ability and resistance to neomycin. The strain could degrade filter paper cellulose, cellulose powder, microcrystalline cellulose, cotton wool, rice straw and gelatin, and it was also able to utilize abundant saccharides as substrates such as cellobiose, glucose, xylose, xylan, raffinose, maltose, sorbose, fructose and galactose. The growth pH ranges from 6.5 to 8.5, temperature from 35 to 70 degrees C and concentration of NaCl on cellulose from 0% to 1.0%, while the optima of pH 6.85, 60 degreesC and 0.2% NaCl. Under the optimal growth conditions, the filter paper cellulose degradation rate was up to 90.40% after 10 days. The optimum temperatures for FPA, CMCase, beta-glucosidase and xylanase were 70 degrees C, 70 degrees C, 70 degrees C, and 60 degrees C respectively. CMCase activity was found with high thermal stability. The phylogenetic analysis based on partial 16S rDNA revealed that HCp was close to Acetivibrio cellulolyticus and A. cellulosolvens with 97.5% sequence similarities. Strain HCp is thermophilic, efficiently cellulolytic anaerobe. It is able to utilize vast substrates and produce highly thermostable enzymes. It is a potential bacterium that can be used for cellulolytic ethanol production.

Han Z.-Y.,Biogas Institute of Ministry of Agriculture | Liu D.,Southwest Jiaotong University | Li Q.-B.,Southwest Jiaotong University
Journal of Environmental Management | Year: 2013

A removing mechanism for organics and nitrogen using a semi-aerobic aged refuse biofilter (SAARB) was evaluated based on the space structure, the aged refuse conformation and characteristics, as well as the degradation theories of organic matter and nitrogen-based substances, which could provide a fundamental theory to more effectively treat organic matter and nitrogen-based pollutants in leachate. The experimental results indicated that the average removal rate of chemical oxygen demand and total nitrogen reached 96.61 and 95.46%, respectively. The aerobic-anoxic-anaerobic zones appeared alternately in both the space structure and the granule conformation inside of the SAARB, which promoted various physical, chemical and biological reactions. Most biodegradable organic matter was converted to CO2 and CH4. The average CO2 release rate was 1.567 L/(h m2) in the winter and 1.467 L/(h m2) in the summer during a single-period experiment. The average CH4 release rate was 0.303 L/(h m2) in the summer; however, it could not be detected in the winter. Moreover, the nitrogen-based pollutants were mostly converted to N2 and N2O through denitrification. Some of the refractory organic matter and nitrogen-based pollutants were likely adsorbed by the aged refuse and biodegraded more slowly. The adsorption rate of biologically degradable matter (BDM) was 0.624 g/(kg d) during the first 40 weeks and the largest absorbance of total nitrogen (TN) was about 7.0 g/kg during this experiment. Therefore, the SAARB can maintain stable and highly efficient environment for removing organic matter and nitrogen-based pollutants. © 2012 Elsevier Ltd.

Liu Q.,Biogas Institute of Ministry of Agriculture
Wei sheng wu xue bao = Acta microbiologica Sinica | Year: 2010

We studied the physiological, biochemical properties and metabolism of Clostridium lituseburense P4-1 from soil in Namucuo. We adopted Hungate anaerobic technique to get Strain P4-1 from soil in Namucuo. Through physiological, biochemical and phylogenetic analysis, we identified the strain P4-1. Cells were Gram-positive and spore-forming. It grew between 13 and 40 degrees C (optimum at 37 degrees C), between pH value 5.0 and 10.O (optimum at 7.5-8.0), and with the presence of NaCl between 0%-5%. Strain P4-1 could metabolize many carbon sources including glucose, melibiose and mannitol. Metabolites of glucose were acetate, butyrate, propionate, CO2, and little H2. Based on 16S rDNA studies, strain P4-1 was most close to Clostridium lituseburense DSM 797 (M59107) with 98.7% similarity. Strain P4-1 could degrade p-toluene sulfonate. Strain P4-1 tolerated low temperature, salt and could degrade p-toluene sulfonate. Its metabolites produced by fermentation of glucose could improve the soil micro-environment. It was significant for strain P4-1 to be utilized in the wastewater treatment.

Zhang W.,Biogas Institute of Ministry of Agriculture
Wei sheng wu xue bao = Acta microbiologica Sinica | Year: 2011

We screened a thermophilic xylolytic bacterium that produced fuel ethanol from a high-temperature oil reservoir, and provided microbial resources to genetic engineering strains construction and consolidated bioprocessing. We adopted Hungate anaerobic technique to isolate strain xyl-d from oil reservoir water sample enriched for two years from Shengli Oilfield in China, and we identified strain xyl-d with morphological, physiological, biochemical and phylogenetic analysis. Strain xyl-d was gram-negative, rod-shaped, spore-forming and strictly anaerobic. The growth temperature ranged from 30 degrees C to 85 degrees C (optimum 65 degrees C) and the pH ranged from 3.0 to 10.0 (optimum 7.5) and salt concentration was 0% - 4% (optimum at 2.0%). It converted D-xylose into ethanol, acetate, CO2, trace amount of iso-butanol and propionate. The genomic DNA G + C contents of strain xyl-d was 45.6 mol%. Based on 16S rRNA gene sequence, strain xyl-d was most close to Thermoanaerobacter wiegelii DSM10319(T) and Thermoanaerobacter ethanolicus DSM 2246(T) both with the 99.3% similarity. It produced more ethanol and less acetate at initial pH 8.5 than other pH. Ethanol yield was increased significantly with yeast extract, and ethanol became the main end product. In addition, growth of strain xyl-d was inhibited obviously with ethanol concentration more than 7% (V/V). In the optimum growth conditions, xylose degradation rates reached to 91.37%. Strain xyl-d was thermophilic, high xylose conversion rate, acidotolerant anaerobe. It was a potential bacterium that can be used for consolidated bioprocessing.

Fan S.,University of Sichuan | Xiao Z.,University of Sichuan | Zhang Y.,University of Sichuan | Tang X.,Biogas Institute of Ministry of Agriculture | And 4 more authors.
Bioresource Technology | Year: 2014

A continuous and closed-circulating fermentation (CCCF) system with a pervaporation membrane bioreactor was built for ethanol fermentation without a refrigeration unit to condense the permeate vapor. Two runs of experiment with a feature of complete and continuous coupling of fermentation and pervaporation were carried out, lasting for 192h and 264h, respectively. The experimental measurement indicated that the enhanced fermentation could be achieved with additional advantages of convenient permeate recovery and energy saving of the process. During the second experiment, the average cell concentration, glucose consumption rate, ethanol productivity, ethanol yield and total ethanol amount produced reached 19.8gL-1, 6.06gL-1h-1, 2.31gL-1h-1, 0.38, and 609.8gL-1, respectively. During the continuous fermentation process, ethanol removal in situ promoted the cell second growth obviously, but the accumulation of the secondary metabolites in the broth became the main inhibitor against the cell growth and fermentation. © 2014 Elsevier Ltd.

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